Development of immobilized beta1-adrenoceptor chromatography for rapid discovery of ligands specifically binding to the receptor from herbal extract
Development of immobilized beta1-adrenoceptor chromatography for rapid discovery of ligands specifically binding to the receptor from herbal extract
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DOI：
10.1016/j.chroma.2022.463298
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发表时间：
2022-07-06
影响因子：
4.1
通讯作者：
Zhao，Xinfeng
中科院分区：
文献类型：
--
作者：
Shayiranbieke，Aerduosi;Liang，Qi;Zhao，Xinfeng
β1-肾上腺素受体 (β1-AR) 配体的发现被视为对抗受体介导的疾病（包括心血管疾病）的巨大需求。由于缺乏高效的铅筛查方法，这种追求受到严重挑战。这项工作开发了一种从草药提取物中寻找β1-AR配体的色谱方法，通过在其C末端融合表皮生长因子受体（EGFR）作为标签，以在E中稳定表达融合受体。大肠杆菌，将表达的EGFR标记的β1-AR固定到依鲁替尼衍生的氨基微球上，并将固定化受体应用于配体-受体相互作用和草药提取物的分析。 X射线光电子能谱和经典药物保留行为等综合表征表明，微球表面修饰的EGFR和依鲁替尼通过共价反应制备的固定化β1-AR具有较高的特异性和良好的稳定性。阿替洛尔、美托洛尔和艾司洛尔的正面分析证实了它们与 β1-AR 的结合，关联常数为 1.07 × 104、6.54 × 103 和 1.45 × 104M−1。热力学分析提供了静电相互作用、氢键和驱动这些相互作用的范德华力的证据。通过反相高效液相色谱-串联质谱法分析保留峰，胡薄荷酮被认为是一种与枣提取物中的β1-AR特异性结合的生物活性化合物。这些结果综合起来表明，当前的方法可以为从复杂基质（如草药提取物）中高效发现 β1-AR 配体提供一种替代方法。
The discovery of beta1-adrenoceptor (β1-AR) ligands is viewed as an enormous demand for fighting ailments mediated by the receptor including cardiovascular diseases. Such pursuit is gravely challenged due to the lack of lead screening methods with high efficiency. This work developed a chromatographic method for pursuing β1-AR ligand from the herbal extract by fusing epidermal growth factor receptor (EGFR) as a tag at its C-terminus to stably express the fusion receptor inE. coli, immobilizing the expressed EGFR-tagged β1-AR onto ibrutinib-derivatized amino microspheres, and applying the immobilized receptor in the analysis of ligand-receptor interaction and herbal extract. Comprehensive characterizations like X-ray photoelectron spectroscopy and retention behaviors of canonical drugs demonstrated high specificity and good stability of the immobilized β1-AR prepared through the covalent reaction between the EGFR and ibrutinib decorated on the microsphere surface. Frontal analysis of atenolol, metoprolol, and esmolol confirmed their bindings to β1-AR with association constants of 1.07 × 104, 6.54 × 103, and 1.45 × 104M−1. The thermodynamic analysis provided proof of electrostatic interaction, hydrogen bonds, and van der Waals force driving those interactions. Pulegone was recognized as a bioactive compound that specifically binding to β1-AR from the extract ofZiziphora clinopodioides Lamby analyzing the retention peak through reverse-phase high performance liquid chromatography coupled with tandem mass spectrometry. These results, taken together, indicated that the current method is possible to provide an alternative for discovering β1-AR ligands with high efficiency from complex matrices like herbal extract.